tbuckley/cabot-search huggingface dataset:
DDDR-18. SINGLE-CELL FUNCTIONAL GENOMIC ANALYSIS OF PHARMACOLOGIC RESPONSES IN <i>NF1</i> MUTANT GLIOBLASTOMA REVEALS DISTINCT MECHANISMS OF ACTION FOR MEK AND SHP2 INHIBITION
Authors: Sixuan Pan, John Liu, Kanish Mirchia, Emily Payne, Joanna Pak, Kyounghee Seo, Tomoko Ozawa, David Raleigh, Mitchel Berger, Adam Abate, Harish Vasudevan
Journal: Neuro-Oncology (2023)

Abstract BACKGROUND Glioblastomas harbor recurrent mutations in NF1 that activate Ras/Raf/MEK signaling, motivating pharmacologic approaches targeting downstream (MEK) or upstream (SHP2) of Ras signaling. However, the mechanisms underlying therapeutic response remain unclear. Here, we integrate genome wide CRISPRi screens and single cell sequencing in cell lines and mouse xenografts to identify functional genomic mechanisms underlying responses to the MEK inhibitor selumetinib or the SHP2 inhibitor RMC-4550. METHODS Human NF1 mutant GBM43 cells expressing CRISPRi machinery were used for CRISPRi screens with selumetinib or RMC-4550. Intracranial xenografts were established with GBM43 cells and treated with selumetinib or RMC-4550. Tumor growth kinetics were measured with bioluminescent imaging. RESULTS Genome-wide CRISPRi screens with selumetinib or RMC-4550 in NF1 mutant human GBM43 cells identified 39 genes (14 sensitivity, 25 resistance) mediating selumetinib response and 327 genes (51 sensitivity, 276 resistance) mediating RMC-4550 response. While both selumetinib and RMC-4550 hits were enriched for components of Ras/Raf/MEK, hits sensitizing RMC-4550 response were uniquely enriched for PI3K signaling and focal adhesion components while hits mediating resistance were enriched for cell cycle genes. In NF1 mutant GBM43, RMC-4550 significantly reduced tumor volume (p=0.02) while selumetinib did not. Single cell RNA-sequencing (scRNA-seq) analysis of 90,782 tumor cells from NF1 mutant GBM43 xenografts treated with selumetinib or RMC-4550 identified 12 clusters across all samples. RMC-4550, but not selumetinib, resulted in significant depletion of cells expressing an epithelial-mesenchymal transition (EMT) signature marked by expression of focal adhesion genes mediating RMC-4550 sensitivity, suggesting this signature comprises a distinct mechanism of action in response to RMC-4550 but not selumetinib. CONCLUSIONS In NF1 mutant glioblastoma in vitro and in vivo models, cellular response to both upstream (SHP2) and downstream (MEK) inhibition are mediated by Ras/Raf/MEK signaling while RMC-4550 alone uniquely regulates a focal adhesion network associated with EMT and demonstrates improved efficacy in vivo.
RADT-19. FACTORS PREDICTIVE OF OUTCOME IN GAMMA KNIFE STEREOTACTIC RADIOSURGERY FOR CEREBRAL METASTASES
Authors: Vassili Crispi, Dharsshini Reveendran, Paul Hatfield, Ryan Mathew
Journal: Neuro-Oncology (2023)

Abstract BACKGROUND Stereotactic radiosurgery (SRS) in brain metastases is associated with prolonged quality of life, locoregional control, and stabilisation of neurological symptoms. Optimal patient selection is pivotal to achieving improved overall survival (OS). This study aimed to evaluate patient and disease factors associated with optimal outcomes. METHODS This retrospective study analysed 3,940 intracranial metastases in 1,113 patients treated with SRS at the Leeds Gamma Knife Centre between 2010 and 2020. Data on primary tumour, Karnofsky performance status (KPS), intracranial disease burden, steroid therapy, and treatment toxicity were recorded. Follow-up MRI imaging was performed at 6 months. OS curves were compared using the Log-Rank test (p &amp;lt; 0.05). RESULTS Multivariate analysis revealed primary diagnosis (p &amp;lt; 0.001), KPS (p &amp;lt; 0.001), number of lesions (p = 0.005) and total tumour volume (p = 0.012) as significant predictors of median OS. Breast carried the highest median OS (18 months) compared to gastrointestinal tumours (7 months) and metastases of unknown origin (5 months). Higher functional performance at baseline was associated with greater median OS: from KPS 100 (18 months) to KPS 70 (6 months). Lower intracranial disease burden conferred greater median OS: patients with 1-3 treated lesions survived significantly longer (11 months) than those with 4 or more (9 months)(p = 0.001). Median OS reduced with an increasing total volume of treated metastases: 13 months if &amp;lt;5000mm3, compared to 4 months if &amp;gt;30000mm3. Following SRS, 375(33.7%) patients had new brain metastases with a median time of 6 months. 13.3% of these were local recurrence, and median time to recurrence ranged from 1.5 months in renal primary to 14.3 and 17.7 months in breast and gastrointestinal tumours, respectively. 195(17.5%) patients underwent further SRS with a median time of 7 months. CONCLUSION Primary diagnosis, KPS, and intracranial disease burden are significant pre-treatment predictors of OS for SRS in brain metastases.
BIOM-36. PREOPERATIVE IMMUNE AND INFLAMMATORY FACTORS IN PERIPHERAL BLOOD PREDICT PROGNOSIS OF GLIOMA AND CORRELATE WITH GRADES AND SUBTYPES: A PROSPECTIVE MULTI-INSTITUTIONAL STUDY OF 1282 PATIENTS
Authors: Sheng Zhong, Wenzhuo Yang, Zhiyun Zhang, Haoqun Xie, Ke Sai, Yonggao Mou
Journal: Neuro-Oncology (2023)

Abstract BACKGROUND Peripheral blood immune and inflammatory factors can reflect glioma patients' immune status and inform anti-tumor immunotherapy, but research in this field is limited compared to tumor microenvironment studies. We aim to investigate the relationship between these circulating factors and glioma grades, subtypes, and prognosis. METHODS From October 2006 to April 2022, 1282 patients with primary gliomas were enrolled in this study, and their preoperative peripheral blood was collected. Correlations between immune and inflammatory factors and different glioma grades and subtypes were analyzed using ANOVA, t-test, and Ordinal Logistic Regression. ROC curve analysis was used to evaluate the diagnostic efficiency of these factors. In addition, the Cox regression model, K-M survival analysis, and other methods were used to study the relationship between these factors and the prognosis and survival of patients. RESULTS The results showed that with the higher clinical grade, the percentage of NK cells in peripheral blood increases while the percentage of Th and T lymphocytes decreases. C4 levels and percentage of B lymphocytes were higher in grade I gliomas than in other grades. The percentage of NK and Th cells also correlated with glioma subtypes. Immunoglobulin light chains were associated with a favorable prognosis, whereas a higher percentage of peripheral B lymphocytes in grade IV patients was associated with a poor prognosis. CONCLUSIONS The relationship between immune and inflammatory factors and the grades and subtypes of glioma can offer potential opportunities to intervene in these factors that could improve the prognosis of glioma patients. Additionally, it may be helpful to screen glioma and anti-tumor immunotherapy.
INNV-19. LANGUAGE ACTIVATION AREAS AND PATHWAYS INTEGRITY ASSESSMENT FOLLOWING GLIOMA RESECTION AND SUBSEQUENT RADIATION TREATMENT
Authors: Ory Haisraely, Zvi Cohen, Wohl Anton, Abigial Livny, Galia Tsarfaty, Eugenia Belenky, Thila Kaisman-Elbaz
Journal: Neuro-Oncology (2023)

Abstract BACKGROUND It is a well-established practice to perform brain-mapping scans of patients diagnosed with dominant frontal/temporal lesions to evaluate their language activation areas (LAAs) and pathways prior to surgical intervention. Dependent on the pathology, brain functional regions and their associated pathways may be exposed to post-operative radiation as part of the patient's treatment protocol. The post-radiation detrimental effect on cognition is well-known, however, the effect of surgical resection and subsequent radiation on LAAs and pathways' integrity is not well studied. Here, the impact of surgical resection and radiation on LAAs and pathways integrity is examined. METHODS 8 patients that underwent surgical resection to frontal/temporal lesion located at their dominant hemisphere followed by radiation treatment with at least two brain-mapping scans in Sheba Medical Center between the years 2012 to 2021 were collected. Location of functional LAAs (such as Broca's and Wernicke’s areas) as well as that of structural fiber tracts (such as the arcuate fasciculus) were determined. Volumetric and tumor geometry data was measured and documented. Laterality index of activation and Fractional Anisotropy (FA) quantitative values were calculated and compared from the baseline scan to the post-treatments scan for each patient. RESULTS Patients’ demographics analysis showed that their median age-at-diagnosis was 41 years (range, 4-64). The median overall survival was 89 months (range, 19-130) and the median follow-up duration was 80 months (range, 16-129). Radiation mean dose was 57Gy (range 40.05-60) and Planning treatment volume was 315ml (range 123.3-456.5) Notably, the mean laterality index of LAAs was also affected by intervention (p=0.044). Additionally, the arcuate fasciculus’ FA values differed in the post-interventions scan compared to the pre-interventions scan CONCLUSION surgical resection and radiation treatment may cause changes in the lateralization of the LAAs that may represent an attempt to compensate for a significant brain injury.
EPCO-17. 3D GENOME STRUCTURE REGULATES TRANSCRIPTION IN PEDIATRIC HIGH-GRADE GLIOMA
Authors: Tina Huang, Juan Wang, Ye Hu, Andrea Piunti, Elizabeth Bartom, Feng Yue, Amanda Saratsis
Journal: Neuro-Oncology (2023)

Abstract INTRODUCTION Pediatric high-grade gliomas (pHGGs), including glioblastoma multiforme (GBM) and diffuse intrinsic pontine glioma (DIPG), are highly morbid brain tumors. Up to 80% of DIPGs harbor a somatic missense mutation in genes encoding Histone H3. To investigate whether the H3K27M mutant protein is associated with distinct chromatin structure affecting transcription regulation, we generated the first high-resolution Hi-C and ATAC-Seq maps of pHGG cell lines, and integrated these with tissue and cell genomic data. METHODS We generated sequencing data from patient-derived cell lines (DIPG n=6, GBM n=3, normal n=2) and frozen tissue specimens (DIPG n=1, normal brainstem n=1). Analyses included cell line RNA-Seq, ChIP-Seq (H3K27ac, H3K27me3, H3K27M) and genome-wide chromatin conformation capture (Hi-C), as well as tissue ATAC-Seq. Publicly available pediatric glioma tissue ChIP-Seq data was integrated with cell data. CRISPR knock-down of target enhancer regions was performed. RESULTS We identified tumor-specific enhancers and regulatory networks for known oncogenes in DIPG and GBM. In DIPG, FOX, SOX, STAT and SMAD families were among top H3K27Ac enriched motifs. Significant differences in Topologically Associating Domains (TADs) and DNA looping were observed at OLIG2 and MYCN in H3K27M mutant DIPG, relative to wild-type GBM and normal cells. Pharmacologic treatment targeting H3K27Ac (BET and Bromodomain inhibition) altered these 3D structures. Functional analysis of differentially enriched enhancers in DIPG implicated SOX2, SUZ12, and TRIM24 as top activated upstream regulators. Distinct genomic structural variations leading to enhancer hijacking and gene co-amplification were identified at A2M, JAG2, and FLRT1. CONCLUSION We show genome structural variations enhancer-promoter interactions that impact gene expression in pHGG in the presence and absence of the H3K27M mutation. Our results imply that tridimensional genome alterations may play a critical role in the pHGG epigenetic landscape and thereby contribute to pediatric gliomagenesis. Further studies examining the impact of the alterations is therefore underway.
BIOS-08. A COMPREHENSIVE ANALYSIS OF METRICS AFFECTING GLIOBLASTOMA RANDOMIZED CONTROLLED TRIALS ROBUSTNESS
Authors: Farrukh Hameed, Sam Sathyamurthi, Omar Sajjad, Maadeha Hassan, Kinji Arikawa, Mamindla Priyadarshini, Yan Wang, Pascal Zinn
Journal: Neuro-Oncology (2023)

Abstract INTRODUCTION Randomized controlled trials (RCTs) are the gold standard for evaluating the effectiveness of oncological interventions. RCTs are influenced by various factors, including trial design, outcome measures, sample size, recruitment, and retention. These factors can impact the robustness of a trial. OBJECTIVE We comprehensively evaluate the characteristics and robustness of RCTs supporting current GBM management by evaluating trial design, treatment effects, reporting strategies, primary outcome description, patients’ loss to follow-up as well as author and patient demographics. METHODS We conducted database searches and all GBM RCTs with significant outcomes published between 2010-2022 were included. Trial characteristics, survival times, the fragility index (FI-minimum number of patients needed to switch to nullify trial outcome) were assessed. Multivariable regression was performed to assess association of trial characteristics with robustness. RESULTS Of the 310 RCTs identified, 95 (30.6%) trials reported overall survival as primary end point, 92 (29.7%) progression free survival and 123 (39.7%) examined safety profiles of drugs. Of these, 178 (57.4%) were published from 2015-2022. The trials had a median power of 80.0% (interquartile range [IQR] 80.0-90.0). The median number of enrolled patients was 46 (IQR 24-108) with a median of 2 (IQR 1-5) loss to follow up. Of these trials, 68 were eligible for FI calculation. The median FI was 4.0 (IQR 2.0-11.50), with 38 trials (55.9%) having an FI &amp;lt; 4 and a third of the trials had FI less than number of patients loss to follow up. After adjusting for co-variates, recently published trials, multiple centers trials and commercial sponsorship was associated with significantly higher FI (p&amp;lt; 0.05). CONCLUSION Trials supporting current GBM management guidelines have alarmingly low robustness. While the robustness of trials has improved over time, future trials must consider additional metrics in their planning.
CTIM-10. A SURGICAL WINDOW-OF-OPPORTUNITY CLINICAL TRIAL EVALUATING THE PCSK9 INHIBITOR EVOLOCUMAB IN PATIENTS WITH GLIOMA
Authors: Kirit Singh, Kelly Hotchkiss, Matthew Foster, Chelsea Railton, Emily Blandford, Elizabeth Thomas, Elizabeth S Miller, David Ashley, Annick Desjardins, Henry Friedman, Margaret O Johnson, Allan Friedman, Steve Keir, James Herndon, John Sampson, Chuan-Yuan Li, Peter Fecci, Anoop Patel, Mustafa Khasraw
Journal: Neuro-Oncology (2023)

Abstract Gliomas evade the immune system by downregulating cell surface expression of major histocompatibility class (MHC)-I. Antagonism of proprotein convertase subtilisin/kexin type 9 (PCSK9) increases MHC-I expression and can enhance checkpoint inhibition in vivo. Evolocumab is an FDA approved monoclonal antibody (mAb) for hyperlipidemia that inhibits PCSK9. However, large constructs such as mAbs typically have limited penetrance across the blood-brain-barrier. We report the preliminary results of a surgical window-of-opportunity study evaluating if Evolocumab administered subcutaneously pre-preoperatively is detectable in tumor tissue from patients with newly diagnosed or recurrent glioma (PESKe; NCT04937413). Twenty contemporaneous control samples are prospectively collected from patients undergoing craniotomy without receiving Evolocumab. Although non-randomized, these specimens provide valuable information for exploratory comparison of the treated and untreated specimens in terms of drug penetrance and MHC-I expression. To date, 24 patients have been enrolled (M: 14, F: 10) with a median age of 46.5. Of these, 5 received pre-procedure subcutaneous Evolocumab (420mg), while the remaining 19 were analyzed as controls. In treated patients, Evolocumab was detectable via mass spectroscopy (LC-MS/MS) in all analyzed cases, including in tumor tissue from both contrast-enhancing and non-contrast-enhancing regions. Ratios of drug in tumor relative to intra-operative blood were markedly higher in samples from contrast-enhancing regions compared to non-contrast enhancing regions (median ratio of tumor to blood of 0.1503 vs 0.0248 respectively). The tumor proteome was also analyzed using LC-MS/MS to quantitate MHC-I and MHC-II proteins. Initial analysis found increased MHC-I (HLA-A, HLA-B, HLA-C) quantitation via mass spectroscopic analysis of the tumor proteome in treated samples compared to contemporaneous untreated controls. No significant adverse events have been reported in those who received pre-procedure Evolocumab. Ongoing analyses will seek to determine whether increased MHC-I quantities in tumor correspond with increased cell surface expression via immunofluorescence and immunohistochemistry.
EOLP-04. EXTRA-NEURAL DISSEMINATION OF MYC-ACTIVATED INFANT MEDULLOBLASTOMA DESPITE MULTIMODAL THERAPY
Authors: Alysia Horbaczewski, Chad Jacobsen, Amy Kelly, Katherine Pearsal, Matthew Ladra, Lindsay Kilburn, David Gass
Journal: Neuro-Oncology (2023)

Abstract Extra neural metastases of infant medulloblastoma portend poor prognosis. Previous reports were in patients receiving treatment with radiation alone or patients with ventriculoperitoneal shunt (VPS). Recently with multimodal therapy, reports of extra-neural medulloblastoma are rarely reported. We report two patients with stage M3 MYC-Activated Infant Medulloblastoma who received aggressive therapy yet developed extra-neural relapses. The first case was a 2-year-old male who underwent gross total resection, which was complicated by hydrocephalus requiring VPS placement. He completed treatment as per ACNS0334 and achieved a near-complete response. Next, he received four cycles of five drug anti-angiogenic therapy followed by 2 cycles of expanded access therapy with I-3F8 antibody. He progressed both in brain and spinal cord, then received palliative craniospinal irradiation with boost. Two months following radiation, he died of disease with significant pancytopenia and hypoechoic nodules throughout the liver, suggestive of disseminated disease. Our second patient is a five-year-old girl, who was diagnosed at age two. She achieved a CR after treatment as per ACNS0334. She then enrolled on a clinical trial with Difluoromethylornithine (DFMO) for three months. On initial disease evaluation, she had progression in brain and spine. She underwent palliative radiation with craniospinal irradiation and boost with concurrent Indoximod on phase 2 clinical trial. Following radiation, she received compassionate use indoximod and temadar for 24 months and remained in CR. Three months after stopping therapy, she progressed to left orbit, pelvis and ribs. She is currently receiving palliative care. MYC-activated medulloblastoma has a very poor outcome and requires multimodal including radiation to achieve remission. Despite aggressive therapy, disease is highly evasive and can spread extracranially to multiple areas including bones, bone marrow, liver, and abdomen. Novel therapies and more exhaustive surveillance regiments need to be developed to limit extracranial dissemination and achieve more durable remissions.
EPCO-49. MYC AMPLIFICATION AT DIAGNOSIS REGULATES THERAPY-INDUCED HYPERMUTATION AT RECURRENT GLIOMA
Authors: Quanhua Mu, Ruichao Chai, Yingxi Yang, Tao Jiang, Jiguang Wang
Journal: Neuro-Oncology (2023)

Abstract Cancer progression and resistance to therapy are driven by clonal evolution. Recent studies have shown that adult diffuse gliomas have divergent clonal dynamics. However, the genomic and epigenomic factors that influence molecular changes after treatment are not well understood. To address this, we analyzed sequencing and clinical data from 544 adult diffuse glioma patients to identify predictors of cancer evolution. Using machine learning methods CELLO2, we predict grade progression and treatment-induced hypermutation based on tumor characteristics and clinical features collected at diagnosis. We found that MYC copy number gain is more prevalent as a founding alteration in the East Asian gliomas and it is the most significant predictor of hypermutation under temozolomide treatment. Further experiments using patient-derived gliomaspheres and established glioma cell lines confirmed the critical role of MYC amplification and pathway activation in hypermutagenesis. We also showed that c-Myc binding to open chromatin and transcriptionally active regions increases the vulnerability of genomic regions to TMZ-induced mutagenesis. This study highlights MYC as an early predictor of cancer evolution and provides a machine learning platform to improve patient management by predicting cancer dynamics.
EXTH-02. A TERT-SPECIFIC TCR-LIKE ANTIBODY CAR T CELL APPROACH FACILITATES KILLING OF GLIOBLASTOMA
Authors: Eric Zhu, Susan Gladney, Julie Ritchey, Alexandra Livingstone, Julie O'Neal, Tanner Johanns
Journal: Neuro-Oncology (2023)

Abstract Glioblastoma (GBM) is a difficult form of brain cancer to treat as GBM cells utilize various strategies to evade host immune responses. Prior studies have shown autologous cytotoxic T cells (CTL) modified to express chimeric antigen receptors (CAR) facilitating host CTL responses against cancers. This study investigates the ability of T-cell receptor (TCR)-like antibody clones against TERT to facilitate specific CAR T cell mediated killing of GBM. Lentiviral vectors were created for five TCR-like antibody CAR constructs recognizing an HLA-A*02:01-restricted TERT-derived antigen. Nur77-GFP Jurkat reporter cell lines and 3 healthy donor-derived CTLs transduced with these vectors were used in flow-based and in vitro killing assays to assess specificity and functionality. K562 cells that did, and did not, express HLA-A*02:01 were targets in the flow-based assay. Flow cytometry captured activation of transduced Jurkat cells via GFP and CD69 upregulation. Three luciferase-transduced GBM cell lines were targets in the cytotoxic killing assay and visualized via bioluminescence imaging (BLI). Increased GFP and CD69 expression was seen on Jurkat cells transduced with clones 3A12 and 3H2 only when co-cultured with TERT antigen and A*02 positive K562 suggesting specific recognition of the TERT:HLA complex. Furthermore, healthy donor-derived CTL transduced with 3A12 and 3H2 CAR constructs demonstrated in vitro killing of tumor cells by BLI. Interestingly, the 3H2 construct only showed cytotoxicity in the presence of exogenous TERT antigen. However, when the heavy and light chains of the 3H2 clone were flipped in the lentiviral construct (3H2F), specificity to the TERT:HLA complex was preserved but the CAR T cells now demonstrated cytotoxicity of GBM without needing exogeneous peptide. TCR-like antibody clones 3A12 and 3H2F specifically recognize an HLA-A*02:01-restricted TERT-derived antigen, facilitating effective in vitro cytotoxicity of GBM cells. Ongoing experiments will evaluate the in vivo activity of these TERT-specific CAR constructs and assess potential off-target toxicity.